Sperm manipulation is widely used in clinical practice and research, using a small-sized tool (e.g., micropipette) to interact with a sperm. For example, in Intracytoplasmic Sperm Injection (ICSI), a medical procedure for in vitro fertilization, a sperm is tapped/immobilized by a micropipette, aspirated into the micropipette, and dispensed from the micropipette into an oocyte (i.e., egg cell).
Conventionally, sperm manipulation is performed manually. An operator controls the motion of a micropipette via a joy stick to physically tap/manipulate the sperm under an optical microscope. Sperm immobilization requires a micropipette to press (tap) the sperm tail against a surface (e.g., the bottom of a Petri dish). As such, sperm immobilization can be a challenging procedure that has stringent skill requirements. Due to the fast movement of a healthy sperm (25 μm/sec), a sperm can move out of the field of view of a microscope quickly. The operator needs to carefully monitor the motion of the sperm by looking into the eye pieces of a microscope and manually move the microscope stage to keep the sperm within the field of view, while simultaneously attempting to move the micropipette to tap the sperm tail (under 1 μm in thickness) for immobilization.
Different from the use of a micropipette for sperm manipulation, EPO Pat. Publ. No. 19803651 discloses a method that uses a pulsed laser to immobilize sperm in culture medium. This method provides an alternative approach for sperm immobilization, but does not automate the sperm immobilization procedure. It also does not enable other operations such as aspiration and dispensation of a sperm. U.S. Pat. No. 7,875,845 discloses a system that uses optical tweezers to move microscopic particles (e.g., sperm) between different regions of the system. PCT Publ. No. WO2010115167 discloses a system that uses dielectrophoresis (DEP) fields for extracting and separating microscopic particles (e.g., sperm).
Assisted analysis of sperm and other biological materials has been of great interest to industry and academia. U.S. Pat. Nos. 7,526,116 and 7,720,272 disclose a system for automated identification of a sperm in an area of interest on a microscope slide. U.S. Pat. No. 7,252,642 and US Pat. Appl. Publ. Nos. 2011149287, 2011147591, 2007245812 and 20040146848 disclose a method for quantifying the total sperm concentration in a sample, a video visualization system, and a displaying means for viewing sperm samples. U.S. Pat. No. 7,521,696 discloses a method for analyzing fluorescently labeled biological material. A user uses fluorescence in images to manually track the biological material. U.S. Pat. Nos. 7,042,639, 7,106,502 and 7,268,939 disclose an imaging system that utilizes image processing techniques to identify, detect, and track microscope specimens, such as biological cells. US Pat. Appl. Publ. No. 2010046823 discloses an automated system and method for visually tracking moving objects (e.g., biological cells).
U.S. Pat. Nos. 4,402,614 and 4,176,953 disclose a method that directs a beam of light at sperm, and measures the reflections of light from the sperm per unit time to measure sperm motility. U.S. Pat. No. 7,838,210 discloses a method for sorting sperm cells. U.S. Pat. Nos. 7,943,384, 7,758,811, and 7,799,569 disclose a system for sorting and classifying sperm cells based on certain characteristics of the sperm such as DNA content. U.S. Pat. No. 4,896,967 and U.S. Pat. No. 4,896,966 disclose an optical system that uses a radiation sensing means and signal processing to extrapolate the motion of sperm cells, bacteria, and particles. U.S. Pat. No. 7,855,078 discloses a flow cytometer that is able to separate sperm based on one or more characteristics. US Pat. Publication No. 2011061472 discloses microfluidic devices for separating sperm and for determining sperm quality of a semen sample.
Several algorithms have been developed in the field of computer-assisted sperm analysis (CASA) to track sperm trajectories, measure sperm velocity, and evaluate sperm energetic (L. Shi, J. Nascimento, C. Chandsawangbhuwana, E. Botvinick, and M. Berns, “An automatic system to study sperm motility and energetics,” Biomed. Microdevices, vol. 10, pp. 573-583, 2008; L. Z. Shi, J. M. Nascimento, M. W. Berns, and E. L. Botvinick, “Computer-based tracking of single sperm,” J. Biomed. Opt., vol. 11, 2006; M. Berezansky, H. Greenspan, D. Cohen-Or, and O. Eitan, “Segmentation and tracking of human sperm cells using spatiotemporal representation and clustering,” Progr. Biomed. Opt. Imaging Proc., vol. 6512, 2007; H. Oku, M. Ishikawa, N. Ogawa, K. Shiba, and M. Yoshida, “How to track spermatozoa using high-speed visual feedback,” in Proc. Annu. Int. Conf. IEEE Eng. Med. Biol. Soc. EMBC, 2008, pp. 125-128; and R. P. Amann and D. F. Katz, “Reflections on CASA after 25 years,” J. Androl., vol. 25, pp. 317-325, 2004).
The aforementioned computer assisted sperm analysis methods, however, are only concerned with visual sperm analysis, and do not enable automated sperm manipulation. In view of the foregoing, what is needed is a system and methods for performing automated sperm manipulation. Automated sperm manipulation overcomes the limitations inherent in manual sperm manipulation. To achieve automated sperm manipulation the following may be required: (1) a system integrating multiple motorized positioning devices and imaging devices; (2) computer vision algorithms for tracking the spatial location of the sperm in real time; and (3) motion control methods for controlling the motorized positioners to manipulate sperm.
Holding pipettes are standard tools used in ICSI for immobilizing a single oocyte. Searching for an oocyte and immobilizing it are time consuming. What is also needed is a device, preferably a disposable device, for immobilizing multiple oocytes and for containing sperm on the same device to significantly facilitate ICSI.
There are several existing patents on vacuum-based cell holding devices. US Pat. Appl. Publ. No. 2007048857 and EPO Pat. Publ. No. 1752221 disclose a cell trapping plate with application of negative pressure suction through trapping holes. Similarly, U.S. Pat. Appl. Publ. No. 20050250197 and U.S. Pat. No. 5,262,128 also disclose cell trapping devices via through-holes and vacuum application. The through holes of the devices of these patent documents are on the device surface.
There are no cell holding devices which are capable of immobilizing multiple oocytes and containing sperm on the same device for ICSI use. What is needed is a device design that is (1) able to effectively trap/release multiple oocytes and hold sperm; (2) able to properly contain cells and liquid medium on the device; and (3) disposable and low cost.